A Critical Role for the cccA Gene Product, Cytochrome c ₂ , in Diverting Electrons from Aerobic Respiration to Denitrification in Neisseria gonorrhoeae

Abstract

Neisseria gonorrhoeae is a microaerophile that, when oxygen availability is limited, supplements aerobic respiration with a truncated denitrification pathway, nitrite reduction to nitrous oxide. We demonstrate that the cccA gene of Neisseria gonorrhoeae strain F62 (accession number NG0292) is expressed, but the product, cytochrome c 2 , accumulates to only low levels. Nevertheless, a cccA mutant reduced nitrite at about half the rate of the parent strain. We previously reported that cytochromes c 4 and c 5 transfer electrons to cytochrome oxidase cbb 3 by two independent pathways and that the CcoP subunit of cytochrome oxidase cbb 3 transfers electrons to nitrite. We show that mutants defective in either cytochrome c 4 or c 5 also reduce nitrite more slowly than the parent. By combining mutations in cccA (⌬c 2 ), cycA (⌬c 4 ), cycB (⌬c 5 ), and ccoP (ccoP-C368A), we demonstrate that cytochrome c 2 is required for electron transfer from cytochrome c 4 via the third heme group of CcoP to the nitrite reductase, AniA, and that cytochrome c 5 transfers electrons to nitrite reductase by an independent pathway. We propose that cytochrome c 2 forms a complex with cytochrome oxidase. If so, the redox state of cytochrome c 2 might regulate electron transfer to nitrite or oxygen. However, our data are more consistent with a mechanism in which cytochrome c 2 and the CcoQ subunit of cytochrome oxidase form alternative complexes that preferentially catalyze nitrite and oxygen reduction, respectively. Comparison with the much simpler electron transfer pathway for nitrite reduction in the meningococcus provides fascinating insights into niche adaptation within the pathogenic neisseriae.

Cytochrome c‐based domain modularity governs genus‐level diversification of electron transfer to dissimilatory nitrite reduction

“…Our findings here differ from those of Hopper and colleagues () who reported that mutants expressing c 5 lacking its C‐terminal heme domain and CcoP with a missense mutation disrupting the functionality of its third heme domain retained the ability to reduce nitrite albeit at reduced levels. It remains to be determined whether this disparity relates to differences in methodologies or the strain backgrounds employed.…”

“…It remains to be determined whether this disparity relates to differences in methodologies or the strain backgrounds employed. Hopper and colleagues () argue for a role for cytochrome c 2 in electron donation to NirK. Interestingly, sequences of c 2 show very high identities for the first 132 amino acids between N. gonorrhoeae and N. meningitidis strains, but there are diverse sequences C‐terminal to this (Finn Erik Aas, Marina Aspholm, unpubl.…”

“…To examine this situation in more detail, we preincubated cells with low levels of nitrite. Thus, Nel was cultured for 1 h without nitrite followed by 1 h incubation with 0.5 mM nitrite before diluting the cultures back to OD 600 = 0.2 and adding 2.5 mM nitrite (Hopper et al, 2013). However, the preincubation with low levels of nitrite did not have any effect on the growth, implying that Nel has an inherently low ratio of NO reductase/nitrite reductase activity.…”

The Escherichia coli CydX Protein Is a Member of the CydAB Cytochrome bd Oxidase Complex and Is Required for Cytochrome bd Oxidase Activity

“…One potential target for interaction could be the CydDC complex, a transmembrane complex that functions as a glutathione transporter and that is known to be required for cytochrome bd oxidase activity (53). This would be potentially analogous to the proposed activity of the 56-amino-acid CcoQ subunit of cytochrome oxidase of Neisseria gonorrhoeae, which may bind multiple oxidase-related proteins (54). A third, potentially more intriguing, possibility is that CydX acts as a regulatory subunit of the CydAB complex that has evolved to modify cytochrome bd oxidase activity in certain species of bacteria.…”

Transposon Mutagenesis Identified Chromosomal and Plasmid Genes Essential for Adaptation of the Marine Bacterium Dinoroseobacter shibae to Anaerobic Conditions

“…Class I cytochrome c molecules are small soluble cytochromes that are needed for electron transfer reactions during denitrification in other bacteria (7). In Neisseria gonorrhoeae, the cccA gene product cytochrome c 2 is essential for the shuttling of electrons toward the denitrification machinery (29). Several other unclassified class I cytochromes of D. shibae (Dshi_0508, Dshi_2868) did not influence anaerobic growth (see Table S1).…”